禽流感DNA疫苗重组质粒组织分布的实时荧光定量PCR方法建立  

Development of real-time fluorescent quantitative PCR for detection biodistribution of avian influenza DNA vaccine

作　　者：熊杰[1,2] 彭广能[1] 王国俊[2] 李俊平[2] 姜永萍[2] 陈化兰[2] 

机构地区：[1]四川农业大学动物医学院,四川雅安625014 [2]中国农业科学院哈尔滨兽医研究所农业部动物流感重点开放实验室/兽医生物技术国家重点实验室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2011年第3期203-207,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：十一五国家科技支撑计划(2006BAD06A05);国家863计划(2006AA10A205)

摘　　要：为研究禽流感病毒(AIV)DNA疫苗重组质粒在组织中的分布情况,本研究以AIV DNA疫苗pCAGGoptiHA 5HA基因为检测对象,分别建立检测AIV DNA疫苗重组质粒的SYBR Green Ⅰ实时定量PCR方法和TaqMan MGB实时定量PCR方法。经优化比较两种方法的特异性、敏感性、重复性和污染率。结果表明,两种方法的标准曲线线性关系均较好,相关系数均达到0.999;最低检测量为42 copies,特异性强;探针法的重复性优于染料法,污染率低于染料法,因此采用TaqMan MGB实时定量PCR方法检测AIV DNA疫苗重组质粒组织分布情况。To detect the DNA vaccine of pCAGGoptiHA5 in immunized chicks,we established the SYBR Green Ⅰ quantitative real-time PCR（QPCR） and TaqMan MGB QPCR methods with the primers and TaqMan MGB probes designed according to the sequence of pCAGGoptiHA5.After compared specificity,sensibility,reproducibility and contamination,we found that the correlation was above 0.999 and the detection sensibility was 42 copies,but the reproducibility and contamination of TaqMan MGB QPCR were better than SYBR Green I QPCR.Therefore,TaqMan MGB QPCR was a better method to study the distribution and persistence of plasmid DNA vaccine pCAGGoptiHA5 in chickens.

关 键 词：禽流感 DNA疫苗 组织分布 实时荧光定量PCR 

分 类 号：S852.65[农业科学—基础兽医学]